Insulin signalling and functional characterisation of human microvascular endothelial cells in heart failure and diabetes

Endothelial cell dysfunction plays a key role in heart failure and diabetic cardiovascular disease. There is extensive heterogeneity among endothelial cells, the characteristics of which depend on the tissue in which they are located. The metabolic characteristics of microvascular endothelial cells from skeletal muscle and adipose tissue remain largely understudied. Endothelial cells were isolated from the skeletal muscle and adipose tissue of four patient groups: those with either heart failure or type II diabetes, a group with both conditions, and a group with neither condition. After incubating the endothelial cells for 4–5 weeks, until passage 2, the identity of the cells was confirmed by their response to stimulation with vascular endothelial growth factor (VEGF). Flow cytometry confirmed further that >98% of the cells were endothelial in origin. The insulin signalling pathway was then explored in these endothelial cells by real-time PCR and Western blotting. Insulin was used to increase the phosphorylation of some protein reacting. Phosphorylated proteins of Akt and eNOS were identified by using specific phospho-Akt (Ser473) and phospho-eNOS (Ser1177) antibodies. Simple qualification and comparative analysis of expression and signalling were investigated. Endothelial cell functional characterisation was explored. Our findings indicated that while diabetes conditions impaired adipose tissue angiogenesis, HF conditions produced the most superoxide generation. A comprehensive understanding of the molecular mechanisms of endothelial cell function may aid in developing more effective therapeutic drugs to improve symptoms and survival rates in heart failure and diabetes patients. The overall aim of this research was to investigate downstream of the insulin signalling pathway of microvascular endothelial cells in these patients.